Inglés Nivel

2
FCEN
READING MATERIAL

2018

Prof. Goldsack
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Unit 5

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UNIT 5

Structure of a Scientific Paper

How Scientists Read
What is the structure of a scientific paper?

All scientific papers have the same general format. They are divided into distinct sections and each
section contains a specific type of information. The number and the headings of sections may vary
among journals, but for the most part a basic structure is maintained. Typically, scientific papers are
comprised of the following parts:

 Title
 Abstract
 Introduction
 Methods
 Results
 Discussion
 Acknowledgments
 Literature cited

Because scientific papers are organized in this way, a reader knows what to expect from each part of
the paper, and they can quickly locate a specific type of information.

Let's examine the content in each section of a scientific paper, and discuss why each section may be
useful to you as a reader.

TITLE. The title will help you to determine if an article is interesting or relevant for your project.
Well-written titles give a reasonably complete description of the study that was conducted, and
sometimes even predict the findings. Included in a title are the species studied, the kinds of
experiments performed, and perhaps a brief indication of the results obtained.

ABSTRACT. Abstracts provide you with a complete, but very succinct summary of the paper. An
abstract contains brief statements of the purpose, methods, results, and conclusions of a study.
Abstracts are often included in article databases, and are usually free to a large audience. Thus, they
may be the most widely read portions of scientific papers.

INTRODUCTION. You will find background information and a statement of the

author's hypothesis in the introduction.
An introduction usually describes the theoretical background, indicates why the work is important,
states a specific research question, and poses a specific hypothesis to be tested.

METHODS. The methods section will help you determine exactly how the authors performed the
experiment.
The methods describe both specific techniques and the overall experimental strategy used by the
scientists. Generally, the methods section does not need to be read in detail. Refer to this section if
you have a specific question about the experimental design.
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RESULTS. The results section contains the data collected during experimentation. The results
section is the heart of a scientific paper. In this section, much of the important information may be in
the form of tables or graphs. When reading this section, do not readily accept an author's statements
about the results. Rather, carefully analyze the raw data in tables and figures to draw your own
conclusions.

DISCUSSION. The discussion section will explain how the authors interpret their data and how
they connect it to other work. Authors often use the discussion to describe what their work suggests
and how it relates to other studies. In this section, authors can anticipate and address any possible
objections to their work. The discussion section is also a place where authors can suggest areas of
improvement for future research.

ACKNOWLEDGMENTS. The acknowledgments tell you what people or institutions (in addition to
the authors) contributed to the work. In reading the acknowledgments, you can see what sources
provided financial support for the study. You might want to know an industry group or the federal
government funded the study.

LITERATURE CITED. This section provides the sources cited throughout the paper.
This section offers information on the range of other studies cited: Does the author cite only his or
her previous studies? Are both classic and modern sources influencing this work? Does the author
look to the work of scientists in other disciplines? The literature cited section is also helpful for
generating a list of background reading on the topic under study.

The Introduction section.

3 main questions are usually addressed in the introduction:

 What is the major theory that frames the work being done?
 What research question is under investigation?
 What specific hypothesis (or hypotheses) is being tested?

To understand how these questions relate to each other, let's look at a classic diagram describing the
production of scientific knowledge.

Two processes are depicted in this diagram:

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  Induction is the development of a general theoretical framework (also termed a paradigm)
based on the available evidence and using both logical reasoning and creative thought. In the
diagram above, we see that inductive thinking can produce theories.
 Deduction is the process of making specific predictions based on a general theoretical
framework and using these predictions to guide the collection of experimental evidence.
Deductive thinking combined with experiments produces evidence

Most biological studies investigate a particular research question that arises within a major theoretical
framework. In such studies, theory and evidence working in close association. To better understand
this, let's consider the research question:

Are athletics detrimental or beneficial?

In the late 1800's and early 1900's, many people began participating in competitive athletics,
including team sports and racing. Many scientists and physicians felt that while moderate exercise
was beneficial, the intense exercise of athletics was detrimental to human health. Many specific facts
supported this view. For example, it was observed that athletes had "swollen" hearts. There were
even recorded incidences of athletes dying while in competition. Thus, a theory that intense exercise
was detrimental began to develop.

Theoretical frameworks allow scientists to develop specific research questions that investigate the
effects of exercise. For example, the mechanism of the development of an enlarged heart in athletes
might be explored. The results of these studies will contribute to the cumulative evidence and may
lead scientists to modify or even overturn the major theory.

How are Hypotheses Developed?

The diagram below describes the scientific process in greater detail. Note the addition of several new
terms.

 Research questions focus on an area where knowledge is incomplete. Specific research

questions are proposed by scientists operating within a particular theoretical framework.
 Hypotheses are tentative explanations that could answer the research question. Scientists use
the prevailing theory as well as the results from particular experiments as a guide in
developing hypotheses.
 Specific predictions are then developed by deductive reasoning. These predictions are based
on a particular hypothesis and should be testable by experiment.
 Finally, experiments are performed to test the specific predictions. The experimental process
will be covered closely in the next lesson.

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This is a very generalized model of the scientific process. All studies are not developed in exactly
this format, but most studies include some of the elements described above.

EXAMPLE - Are athletics detrimental or beneficial? (continued)

Let's return to our example. Scientists working within the theory that athletics are detrimental might
ask research questions relating to the swollen heart of athletes. Here's how that might work:

 Research question: How do athletics impact cardiovascular health?

 Hypothesis: The enlarged heart of athletes contributes to health problems.
o Notice that this hypothesis is developed by combining the overall theoretical
framework (athletics are bad) with some specific experimental results (athletes have
enlarged hearts).
 Specific prediction: Athletes have a greater risk of heart attacks than the general population.
o Notice that this prediction can be directly tested by experiment.
 Experiment: The rate of heart attacks is measured in athletes and non-athletes.

Note that the scientific process is a continuous process. The specific result obtained from an
experiment can be used to modify the hypothesis. For example, scientists might refine their initial
hypothesis to state "The enlarged heart of elderly athletes contributes to health problems." Again,
specific predictions could be made and these predictions tested by experiment.

If experimental testing repeatedly does not support the hypotheses that are developed within the
theoretical framework, then the overall theoretical framework may be questioned. Scientists might
then begin to develop a new theoretical framework.

In the example of athletics, the theory that athletics are detrimental to health was not supported by
experimental evidence. Thus, a new theoretical construct was developed. Today, the general theory
about exercise and health is that exercise is required for proper health. Exercise scientists develop
hypotheses using this theory as a guide.

Reflection

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As we have seen, the development of hypotheses is often influenced by the prevailing theoretical
framework. Starting with the observation that athletes often have enlarged hearts, let's consider how
scientists with different theoretical frameworks might proceed.

 Imagine that a scientist believes athletics are detrimental to overall health. What sorts of
hypotheses might this scientist generate to explain the enlarged heart of athletes?
 Now imagine that another scientist believes that athletics are beneficial to overall health.
What sorts of hypotheses might this scientist generate to explain the enlarged heart of
athletes?
 How do you think that the theoretical framework of these scientists will influence the kind of
studies that they perform?

The Methods section

In the methods section, scientists provide specific information on how the hypothesis was tested. It is
important that the methods section is detailed enough for a competent reader to repeat the study and
reproduce its results. This section usually covers four main topics:

 Equipment and materials

 Observations and measurements
 Methods of data analysis
 Experimental design

Equipment and materials. The method section provides a description of the equipment and
materials used during experimentation. For example, scientists may note solution concentrations,
specific research techniques or sampling procedures, as well as model and catalog numbers of
equipment used during experimentation. Scientists generally provide the species name and a brief
description of any living organism used in their experiment.

Observations and measurements. Experimental measurements and observations are also reported in
the methods section. Here, scientists mention any unexpected developments or observations. They
also explain what data was collected and how and when their measurements were obtained. Imagine
that a scientist is investigating the influence of exercise on blood osmolality. Their data collection
would be described precisely in the methods section and would read something like this: blood
osmolality was measured at 1, 2, and 3 hrs after exercise using a vapor pressure osmometer.

Methods of data analysis. Experimental data are generally evaluated using statistical tests and
computer programs. The methods section usually specifies the type of statistical analysis used to
assess data. The results are not discussed at this point in the paper.

Experimental design. Scientists also use the methods section to explain the strategy of their study.
This strategy is known as the "experimental design" and is the foundation of any scientific
investigation. Because experimental design is so important to the success and validity of a scientific
paper, we will discuss it in greater detail on the next page.

Experimental design

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Experienced readers critically evaluate experimental design before drawing conclusions about a
study.

Study design

The reliability of a study is largely determined by the overall experimental design.

A case study intensely describes one subject or a small group of subjects. Scientists performing case
studies can only draw conclusions about the subjects examined. Case studies often lead to the
formation of hypotheses that can be tested by more rigorous experimental designs.

A cross-sectional study is the observation of a defined population at a single point in time. Scientists
do not actively manipulate experimental variables in this design, rather they look for connections
among variables. Cross-sectional studies are useful for establishing correlation between two
variables, but they usually do not establish causation.

A longitudinal study involves data collection over a defined period of time. Scientists establish
experimental treatments and examine the effects of changing a particular variable. This design allows
the researcher to measure changes in variables over time in response to an experimental treatment
and establish causation.

Example: The different types of research design

Research question: Do low carbohydrate diets promote weight loss?

Case study: A scientist places two overweight patients on a low carbohydrate diet and measures
weight. If these patients lose weight, there is suggestive evidence that a low carbohydrate diet may
contribute to weight loss in some individuals.

Cross-sectional study: A scientist records the diets and weights of 500 individuals. The weights of
those who eat a low carbohydrate diet are compared to those eat other diets. This study could
establish a correlation between low carbohydrate diets and lower body weights. However, it can not
establish that low carbohydrate diets cause lower body weights, because many factors have not been
controlled in this study design.

Longitudinal study: A scientist places 20 subjects on a low-carbohydrate diet and compares their
weight to 20 control subjects on a normal diet. Because this study establishes an experimental
treatment (the low carbohydrate diet) and controls other variables, it could support the hypothesis that
low carbohydrate diets cause lower body weights.

Sample size

The sample size is the number of subjects or individuals studied. In scientific literature, the letter "N"
(or "n") is used to designate sample size. In experiments, scientists usually work with a sample of
subjects rather than the full population. Good experimental design must include a sample of subjects
that is representative of the greater population being studied.

Control groups

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Scientific hypotheses are tested by contrasting a "control group" with an "experimental group". In
experimental groups, the variable under study is manipulated. In control groups, the variable under
study is not manipulated. Control groups are important because they serve as a reference point for the
experimental groups. In some studies, subjects are studied twice, once with the experimental
manipulation, and once with a control manipulation. In this case, the subject's control measurements
can be compared to their own experimental measurements.

Scientists examining human subjects often use placebos in their control treatments. A placebo is a
substance or treatment that replicates the experimental treatment but lacks the active component.
Placebos give the control subjects the same expectation as the experimental subjects and thus enable
scientists to control for psychological effects.

Replicates

Scientists often need to replicate an experiment several times to establish a clear relationship
between control and experimental groups. Each performance of the experiment is called a replicate.
The number of replicates that a scientist chooses to perform is important; too few replicates could
lead to inconclusive results while too many replicates is a waste of resources.

Reflection
Companies often use testimonials to promote their products. Suppose that you encountered the
following testimonials for a new weight-loss pill.

From a movie star, "I lost 20 pounds without dieting using these pills".

From a research scientist at Your State University: "Our research shows that subjects taking these
pills lose an average of 12 pounds in 4 weeks".

From a professional baseball player, "I've tried all of the diet pills, and these are the only ones that
have worked for me."

From a scientist at the pharmaceutical company that makes the pills, "This is the most advance
weight loss formula on the market".

 Which of these testimonials is most convincing?

 How do testimonials differ from the experimental
designs discussed in this lesson?
 Can you translate these testimonials
into a testable hypothesis?
 What experimental design would effectively test
the hypothesis you describe?

The Results section

Authors present their experimental data in the results section. Often, these data appear in the form of
graphs and tables. Careful scrutiny of the actual experimental data is vital to successfully and
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critically analyzing a scientific paper. Thus, when reading a scientific paper, you should pay close
attention to its graphs and tables. To maintain a critical stance, you may want to examine the graphs
first and then read the text of the results section to see how the authors interpret their data. At the
very least, you will want to verify any important claims the authors make by examining their graphs
and tables.

Reading a graph requires organization and attention to detail. For a successful critique of the data,
you should examine both graphs and figure legends. Figure legends are blocks of text that are usually
located below the graph they describe. Sometimes data presented in a graph can be clarified by
referring to the text of a paper (especially the methods section.)

Let's consider the graph below. (Note: The data for this graph was created to illustrate graph-reading
strategies. The graph does not necessarily demonstrate the actual heart rate responses of men and
women.)

Tips on how to read a graph.

 To understand Figure 1, first examine its X and Y axes (see above). The dependent
variable (the measured variable) is plotted on the Y-axis. The independent variable is
plotted on the X-axis. The independent variable may be a category or attribute (i.e. sex or
species) or it may be an experimentally manipulated factor (i.e. time or exercise intensity).
 Pay careful attention to the units of measurement. 60 beats per minute (bpm) is very different
than 60 beats per second (bps).
 Look for indicators of experimental variability. These are typically found as error bars on a
figure and they are explained in the figure legend. In the example above, the error bars
represent a measure of variability called the standard error. The figure legend reports
this:Values are mean +/- standard error.

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  Determine the number of individual measurements that were made. In the graph above, the
heart rates of 12 men and 12 women were studied. This is indicated by the statement: N=12
for each treatment.
 Look for indications of statistical significance. In the above graph, the asterisk over the
female bar indicates that female heart rates differ significantly from male heart rates. The
statistical tests used to determine significance (in this case, a Student's t-test) are always
identified in a graph's figure legend. The statement p<0.05 means that there is less than a 5%
probability that an indicated difference is due to chance.

EXAMPLE 1 - Interpreting Figure 1.

Now, we are ready to actually interpret a graph. Let's look at Figure 1 (above). Take a moment to
read it. Answer the following questions and then check your answers.

Questions:

1. What is the average resting heart rate for humans?

2. How do the heart rates of men and women differ?
3. How much variability is there in heart rate?

Answers:

1. Between 60 and 70 bpm.

2. Women have slightly higher heart rates than men; this difference is statistically significant.
3. There is some variability in resting heart rate among individuals; the variability is somewhat
higher in women than in men.

Results - Constructing graphs

Authors must make certain decisions when constructing a graph to represent the results of their study.
Consider Figures 2 and 3 below. Both figures use the exact same data set to describe an experiment
with two independent variables: gender and exercise. Notice, however, that each figure stresses
different parts of the experiment.

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(Note: The data for these graphs were created to illustrate graph-reading strategies. The graphs do not
necessarily demonstrate the actual heart rate responses of men and women.)

EXAMPLE 2. Interpreting Figures 2 and 3.

Questions:

1. Based on Fig 2 or 3, does exercise affect heart rate?

2. Based on Fig 2 or 3, is there a gender difference in heart rate?
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 3. Based on Fig 2 or 3, does exercise influence the difference in heart rates between males
and females?
4. Which figure is more effective at stressing a difference in heart rate between males and
females?
5. Which figure is more effective at establishing a difference between exercise and resting?

Answers:

1. Yes, heart rate increases after exercise

2. Yes, females tend to have higher heart rates at rest than males.
3. Yes. In the resting condition, females have a higher heart rate than men. However, after
exercise, the heart rates of males and females do not differ.
4. Figure 2 shows these differences most clearly because it directly compares the heart rates of
males and females based on exercise type.
5. Because heart rates at rest and heart rates during exercise vary greatly, both figures effectively
establish a difference. The magnitude of the difference in exercising and resting heart rates for
each gender can be most easily viewed in Figure 3.

Reflection
Imagine that the resting heart rates of a group of college students were measured (in beats per
minute) and reported in the chart below.

Smokers Non-smokers
56 (b.p.m.) 52
67 61
59 70
87 54
76 63
65 72
72 60
64 64
71 54
78 51

 Based on these data, is there a difference in resting heart rate between smokers and non-
smokers?
 What additional analyses would you like to perform in order to better understand these data?
 What would be the best way to report these data in a scientific paper?

10 Tips for Reading Primary Literature

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 Here are ten tips to guide your reading of the primary literature:

1. Focus on methods and results. Try not to be influenced by the way the study is presented, but
rather focus your analysis on the experimental design, techniques, and data.
2. Be a skeptic. Ask yourself how strongly the authors' interpretations and conclusions are
supported by the evidence.
3. Be fair. Scientific research is difficult, and scientists operate under many constraints. Don't
expect studies to be perfect.
4. Read non-linearly. Exploit the format of research articles to quickly access the information
you need. Don't feel compelled to read every line start to finish. Skim the paper to understand
its overall approach. Refer to previous sections as necessary.
5. Consider the big picture. Assess where the study fits into the cycle of science, and how it
relates to previous research.
6. Consult other sources. Writers of research articles assume their audience has basic knowledge
of the area. Consult secondary sources to get the needed background.
7. Take your time. Research articles condense entire studies into a few printed pages. It probably
took the authors years to conceive, perform, and publish their work. Be patient and persistent
when reading articles.
8. Accept uncertainty. Research articles deal with emerging knowledge and controversial issues.
Don't expect to find absolute answers to every question. Each paper is a step in an ongoing
process.
9. Expect to be challenged. If you're not an expert in an area, there might be aspects of a paper
you can't understand fully. That's OK, you can still learn from those parts of a paper that you
can comprehend.
10. Relax and enjoy. Perhaps this is the hardest advice to follow, especially when you're
confronted with a complicated paper. But try to approach an article like a puzzle. It's going to
take time and effort to make progress, but there's real satisfaction in doing so.

From: Gillen, C.M. Reading primary literature: a practical guide to evaluating research articles in
biology. Benjamin Cummings, San Francisco. 44p., 2007.

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*******************************************************************************

Scientific Communication: Understanding Scientific Journals and Articles

by Anthony Carpi, Ph.D., Anne E. Egger, Ph.D., Natalie H. Kuldell

 Scientists make their research available to the community by publishing it in scientific

journals.
 In scientific papers, scientists explain the research that they are building on, their research
methods, data and data analysis techniques, and their interpretation of the data.
 Understanding how to read scientific papers is a critical skill for scientists and students of
science.

Scientists publish their original research in scientific journals, which are fundamentally different from
news magazines. The articles in scientific journals are not written by journalists – they are written by
scientists. Scientists write articles to describe their findings to the community in a transparent
manner. Within a scientific article, scientists present their research questions, the methods by which
the question was approached, and the results they achieved using those methods. In addition, they
present their analysis of the data and describe some of the interpretations and implications of their
work. Because these articles report new work for the first time, they are called primary literature. In
contrast, articles or news stories that review or report on scientific research already published
elsewhere are referred to as secondary.

The articles in scientific journals are different from news articles in another way – they must undergo
a process called peer review in which other scientists (the professional peers of the authors) evaluate
the quality and merit of research before recommending whether or not it should be published.

SCIENTIFIC JOURNALS

There are thousands of scientific journals that publish research articles. These journals are diverse
and can be distinguished according to their field of specialization. Among the most broadly targeted
and competitive are journals like Cell, the New England Journal of Medicine (NEJM), Nature, and
Science that all publish a wide variety of research articles. Cell focuses on all areas of biology,
NEJM on medicine, and both Science and Nature publish articles in all areas of science. Scientists
submit manuscripts for publication in these journals when they feel their work deserves the broadest
possible audience.

All of these journals play a critical role in the advancement of science and dissemination of
information. However, to understand how science is disseminated through these journals, you must
first understand how the articles themselves are formatted and what information they contain. There
are broad characteristics that all scientific journal articles share:

FORMAT FOR THE PAPER

Scientific research articles provide a method for scientists to communicate with other scientists
about the results of their research. A standard format is used for these articles, in which the author
presents the research in an orderly, logical manner. This format is:

| Title | Authors | Introduction | Materials and Methods | Results (with Tables and Figures)
| Discussion | Acknowledgments | Literature Cited |

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EVALUATING A PAPER

A thorough understanding and evaluation of a paper involves answering several questions:

a. What questions does the paper address?
b. What are the main conclusions of the paper?
c. What evidence supports those conclusions?
d. Do the data actually support the conclusions?
e. What is the quality of the evidence?
f. Why are the conclusions important?

a. What questions does the paper address?

Before addressing this question, we need to be aware that the research can be of several different
types:

Type of research Question asked:

What is there? What do we see?

Descriptive
How does it compare to other organisms? Are our
Comparative
findings general?

Analytical How does it work? What is the mechanism?

Descriptive research often takes place in the early stages of our understanding of a system. We
can't formulate hypotheses about how a system works, or what its interconnections are, until we
know what is there.

Comparative research often takes place when we are asking how general a finding is. Is it specific
to my particular organism, or is it broadly applicable?

Analytical research generally takes place when we know enough to begin formulating hypotheses
about how a system works, about how the parts are interconnected, and what the causal
connections are. A typical analytical approach would be to devise two (or more) alternative
hypotheses about how a system operates. Being aware that not all papers have the same approach
can orient you towards recognizing the major questions that a paper addresses.

Identifying structure
Five common types of structure used in scientific texts are:

 Generalization: the extension or clarification of main ideas through explanations or examples

 Enumeration: listing of facts
 Sequence: a connecting series of events or steps
 Classification: grouping items into classes
 Comparison / contrast: examining the relationships between two or more things

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ANALYSING A SCIENTIFIC PAPER Prof. Balliro

Question Guide

 What is the title of the paper?

 Who is the author?
 Has the work been published? Where? When?
 What is the structure of the paper?
 What is the Purpose/s of publication?
 What audience may be interested in reading this paper?
 Refer to the contents of the abstract
 What is the major theory that frames the work being done?
 What research question is under investigation?
 What specific hypothesis (or hypotheses) is being tested?
 What relevant information for the rest of the work is included in the introduction section?
 Define the part of the paper in which there are tables, graphs or diagrams.
 Is the methods section detailed enough for a competent reader to repeat the study and
reproduce its results? Is there any reference to:
- Equipment and materials

- Observations and measurements

- Methods of data analysis

- Experimental design

 Define the part or section in which the author makes a description, analyses or makes a
comparison
 Are there abbreviations? Are they defined as being used by the first time?
 How is the experimental data presented in the Results section?
 Has the author included any of these items in the discussion section?:
(1) tested hypotheses,

(2) limitations,

(3) similar results of other authors,

(4) predictions that follow the result. Does the discussion include conclusions or
recommendations?

 State the way the citations were made.

 What are the main conclusions of the paper?
 What evidence supports those conclusions?
 Do the data actually support the conclusions?
 What is the quality of the evidence?
 Why are the conclusions important?

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EXAMPLE OF PAPER:

Performance characteristics of biodistribution of 99mTc-

cefprozil for in vivo infection imaging
Mahmoud Hamdi Sanad1* and EH Borai2

 * Corresponding author: Mahmoud H Sanad msanad74@yahoo.com

Author Affiliations
1
Labeled Compounds Department, Hot Labs Center, Atomic Energy Authority, Cairo, Egypt
2
Analytical Chemistry Department, Hot Labs Center, Atomic Energy Authority, Cairo, Egypt

For all author emails, please log on.

Journal of Analytical Science and Technology 2014, 5:32 doi:10.1186/s40543-014-0032-3

The electronic version of this article is the complete one and can be found online at: http://www.jast-
journal.com/content/5/1/32

© 2014 Sanad and Borai; licensee Springer.

This is an Open Access article distributed under the terms of the Creative Commons Attribution
License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution,
and reproduction in any medium, provided the original work is properly cited.

Abstract

Background

Cefprozil, an antibiotic used to treat bacterial infections, was labeled with 99mTc. 99mTc-cefprozil was
prepared by adding 99mTc to cefprozil in the presence of 50 µg SnCl2.dihydrite at pH 4 and 30-
minutes reaction time. The radiochemical yield and purity of 99mTc-cefprozil were evaluated after
species separation individually followed by subsequent quantification of the free and complexed
species.

Methods

4 mg of cefprozil was accurately weighed and transferred to an evacuated penicillin vial. Exactly 50
µg SnCl2 solution was added and the pH of the mixture was adjusted to 4 using 0.1N HCl, then the
volume of the mixture was adjusted to one ml with N2-purged bidistilled water. One ml of freshly
eluted 99mTcO4- (200- 400MBq) was added to the above mixture.
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Results

The obtained maximum radiochemical yield was found to be 97.5% ± 0.8%. The 99mTc-cefprozil
complex was stable for 6 hrs. The biological distribution of 99mTc-cefprozil was experimentally
investigated in induced-infection mice, in the left thigh, using Staphylococcus aureus.

Conclusions

T/NT for 99mTc-cefprozil was found to be 5.5±0.12 at 2 hrs after intravenous injection, which was
higher than that of the commercially available 99mTc-ciprofloxacin followed by gradual decline. The
abscess to normal muscle ratio indicated that 99mTc-cefprozil could be used for infection imaging.
Moreover, 99mTc-cefprozil could efficiently differentiate between bacterial infection and sterile
inflammation.

Keywords:

Cefprozil; Ciprofloxacin; Technetium-99m; Infection; Inflammation

Background

Nuclear medicine scintigraphy (NMS) techniques are proving valuable in understanding the disease
processes at molecular levels without the inconvenient invasive procedures, if a specific ligand is
provided for a pathological process. The role of NMS in the diagnosis of infection and its
discrimination from non-infective inflammatory processes and tumors at an early stage is clinically
vital for appropriate management (Wouter et al. [2010]; Sandip et al. [2009]; David-Axel et al.
[2006]; Carrino et al. [2006]). The radiolabeled leucocytes can be considered as gold standard for the
identification of inflammatory foci in the intestine and bone tissues (Weiner [1990]). The
development of new radiopharmaceuticals that do not require the manipulation of blood and are able
to differentiate between inflammatory and infectious processes with high sensibility and specificity is
considered as the object of recent research. Therefore, other preparations such as 99mTc-nanocolloid,
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Ga-citrate, and 99mTc or 111In-labeled human polyclonal immunoglobulin are currently being tested
(Buscombe et al. [1991]). However, none of the preparations is capable of distinguishing between
infections and inflammatory lesions in a clinically useful manner. Recently, a new proposal is based
on the use of radiolabeled antibiotics. One of the most important radiopharmaceuticals which are now
currently available for imaging infection, the antimicrobial agent ciprofloxacin labeled with 99mTc,
has probably shown significant benefits. The fundamental problems of 99mTc-ciprofloxacin
preparation discussed in the literature (Mcafee et al. [1991]; (Akhtar et al. [2005,2004]; Nibbering et
al. [2004]; El-Ghany et al. [2007]) are related to radiochemical purity as well as to the stability of the
labeled complex. So, other antimicrobial agents such as levofloxacin (El-Ghany et al. [2005]),
pefloxacin (Motaleb [2007a,b]), lomefloxacin (Motaleb [2007a,b]), cefoprazone (Motaleb [2007a,b]),
cefuroxime (Yurt Lambrecht et al. [2008]), rifampicin (Syed et al. [2010]), and amoxicillin (Motaleb
and Sanad [2012]) were labeled with 99mTc to be used for imaging sites of infection and to overcome
the drawback of 99mTc-ciprofloxacin (Kleisner et al. [2002]; Yang et al. [2009]). Cefprozil is a semi-
synthetic broad-spectrum cephalosporin antibiotic. Cefprozil is a cis and trans isomeric mixture
(≥90% cis). The chemical name of cefprozil is (6R,7R)-7-[(R)-2-amino-2-(p-hydroxyphenyl)
19
acetamido]-8-oxo-3-propenyl-5-thia-1-azabicyclo [4.2.0] oct-2-ene-2-carboxylic acid (Petropoulos et
al. [2009]; Shaikh et al. [2008]). Cefprozil has in vitro activity against a broad range of gram-positive
and gram-negative bacteria. The bactericidal action of cefprozil results from inhibition of cell wall
synthesis (Breier et al. [2002]; Can [2011]). In this paper, cefprozil was labeled with the most widely
used imaging radionuclide, 99mTc. Factors affecting the labeling yield of 99mTc-cefprozil complex and
biological distribution in inflammation bearing animals were studied in detail.

Experimental

Chemicals and materials

Cefprozil was purchased from Sigma-Aldrich Chemical Company (St. Louis, MO, USA); its
structure is presented in Figure 1. All the other chemicals were purchased from Merck (Whitehouse
Station, NJ, USA), and they were reactive grade reagents. The water used is purged with nitrogen gas
to give deoxygenated bidistilled water.

Figure 1. The chemical structure of cefprozil.

Apparatus

Well-type γ-scintillation counter: Scalar Ratemeter SR7 (Nuclear Enterprises Ltd., USA); pH meter:
model 601, a digital ion analyzer (Orion Research, Jacksonville, FL, USA); ionization chamber:
model CRC-15R (Capintec, Ramsey, NJ, USA); precision electronic balance: model HA 120 (MAD
Company Ltd., Japan); stirring hot plate: model 210T Thrmix (Fisher, Waltham, MA, USA);
electrophoresis apparatus: E.C. Corporation (Albany, OR, USA)

Labeling of cefprozil

Cefprozil (4 mg) was accurately weighed and transferred to an evacuated penicillin vial. Exactly 50-
μg SnCl2 solution was added, and the pH of the mixture was adjusted to 4 using 0.1 N HCl; then, the
volume of the mixture was adjusted to 1 ml with N2-purged bidistilled water. One millimeter of
freshly eluted (200 to 400 MBq) was added to the above mixture. The reaction mixture was
vigorously shaken and allowed to react at room temperature for a sufficient time to complete the
reaction. This experiment was conducted to study the different factors that affect the labeling yield
such as tin content (as SnCl2·2H2O), substrate content, pH of the reaction medium, and reaction time.
For labeling process, trials were performed for each factor under investigation to obtain the optimum

20
value. The experiment was repeated with all factors kept at optimum values except the factor under
study, until the overall optimal conditions are achieved.

Separation methods and recovery of 99mTc-cefprozil complex

Paper chromatography

The radiochemical yield of 99mTc-cefprozil was determined by paper chromatography, in which the
reaction product was spotted on ascending paper chromatography strips (10 × 1.5 cm). Free in the
preparation was determined using acetone as the mobile phase. Reduced hydrolyzed technetium was
determined by using ethanol/water/ammonium hydroxide mixture (2:5:1) or 5 N NaOH as the mobile
phase. After complete development, the strips were dried, cut into 0.5-cm pieces, and counted in a
well-type γ-scintillation counter.

Electrophoresis conditions

Electrophoresis was done with EC-3000 p-series programmable power and chamber supply units
(E.C. Apparatus Corporation) using cellulose acetate strips. The strips were moistened with 0.05 M
phosphate buffer pH 7.2 ± 0.2 and then were introduced in the chamber. The samples (5 μl) were
applied at a distance of 10 cm from the cathode. The radioactivity values were evaluated at an
applied voltage of 300 V and standing time of 1.5 h. The developed strips were dried and cut into 1-
cm segments and counted by a well-type NaI scintillation counter. The radiochemical yield was
calculated as the ratio of the radioactivity of the labeled product to the total radioactivity

HPLC analysis

A simple, short, rapid, sensitive, and robust reversed phase high-performance liquid chromatographic
(HPLC) method was developed and validated to measure the amount of cefprozil in dissolution
profile. An isocratic elution of filtered sample was performed on cosmosil RP18 LiChrosorb column
(250 mm × 4 mm, 5 μm, Merck) using water/acetonitrile solution (90:10, v/v) as mobile phase and at
a UV detection at 200 nm. The mobile phase was delivered at a flow of 1.0 ml/min and at a
maintained column temperature at 50°C, and quantification was achieved with reference to the
external standards (Asikoglu et al. [2000]). Then, fractions of 1.0 ml were collected separately using
a fraction collector up to 15 ml and counted in a well-type γ-scintillation counter.

Stability of 99mTc-cefprozil in human serum

The stability of 99mTc-cefprozil was studied in vitro by mixing 1.8 ml of normal human serum and
0.2 ml of 99mTc-cefprozil and incubated at 37°C for 24 h. Exactly 0.2-ml aliquots were withdrawn
during the incubation at different time intervals up to 24 h and subjected to paper chromatography for
the determination of the percentage of 99mTc-cefprozil, reduced hydrolyzed technetium, and free
pertechnetate.

Induction of infectious foci

21
A single clinical isolation of Staphylococcus aureus (Boyd [1986]) from biological samples was used
to produce focal infection. Individual colonies were diluted in order to obtain turbid suspension.
Groups of five mice, weighing approximately 25 g each, were injected with 200 μl of the above
suspension in the left thigh muscle. Twenty-four hours was required to get gross swelling in the
infected thighs.

Induction of non-infected inflammation

Sterile inflammation was induced by injecting 200 μl of turpentine oil (Robbins [1984]), which was
sterilized by autoclaving at 121°C for 20 min, intramuscularly in the left lateral thigh muscle of the
Albino mice. Two days later, swelling appeared.

In vitro binding of 99mTc-cefprozil to bacteria

Binding of 99mTc-cefprozil to S. aureus bacteria was assessed by the method described elsewhere
(Welling et al. [2000]). Briefly, 0.1 ml of sodium phosphate buffer containing about 5 MBq of 99mTc-
cefprozil was transferred to a test tube. Exactly 0.8 ml of 50% (v/v) of 0.01 M acetic acid in
phosphate buffer containing approximately 1 × 108 viable bacteria was added. The mixture was
incubated for 1 h at 4°C and then centrifuged for 5 min at 2,000 rpm at 4°C. Simultaneously, the
incubation was performed in the presence of an excess of unlabeled cefprozil (10-, 50-, 100-fold).
The supernatant was removed, and the bacterial pellet was gently resuspended in 1.0 ml of ice-cooled
phosphate buffer and recentrifuged. The supernatant was removed, and the radioactivity in the
bacterial pellet was determined by a γ-counter. The radioactivity related to the bacteria was expressed
in percentages of the added 99mTc activity bound to viable bacteria in regard to the total 99mTc
activity.

Animal studies

The study was approved by the animal ethics committee of the Labeled Compound Department and
was in accordance with the guidelines set out by the Egyptian Atomic Energy Authority. For the
infection model, the animals, Swiss Albino mice (25 to 30 g), were intravenously injected with
100 μl (100 to 150 MBq) of sterile 99mTc-cefprozil, adjusted to physiological pH via the tail vein, and
kept alive in metabolic cage for different intervals of time under normal conditions. For quantitative
determination of organ distribution, five mice were used for each experiment, and the mice were
sacrificed at different times post-injection. Samples of fresh blood, bone, and muscles were collected
in pre-weighed vials and counted. The different organs were removed, counted, and compared to a
standard solution of the labeled cefprozil. The average percent values of the administrated dose/organ
were calculated. The blood, bone, and muscles were assumed to be 7%, 10%, and 40%, respectively,
of the total body weight (Motaleb [2001]). Corrections were made for background radiation and
physical decay during experiment. Both target and non-target thighs were dissected and counted.
Differences in the data were evaluated with Student t test. Results for p using the two-tailed test were
reported, and all the results are given as mean ± SEM. The level of significance was set at P < 0.05.

Results and discussion

22
Separation characteristics

In case of ascending paper chromatographic method, acetone was used as the developing solvent,
free moved with the solvent front (Rf = 1), while 99mTc-cefprozil and reduced hydrolyzed technetium
remained at the point of spotting. In the case of ascending paper chromatographic method, the
mixture was used as the developing solvent; reduced hydrolyzed technetium remains at the origin
(Rf = 0), while the other species migrate with the solvent front (Rf = 1). The radiochemical purity was
determined by subtracting the sum of the percentage of reduced hydrolyzed technetium and free
pertechnetate from 100%.

The paper electrophoresis pattern revealed that the 99mTc-cefprozil complex moved towards the
anode, indicating the anionic nature of this complex. Under similar condition, moved considerably
toward the anode, suggesting that it has a completely ionized negative charge.

An HPLC chromatogram was presented in Figure 2 and showed two peaks, one at fraction no. 3.25,
which corresponds to , while the second peak was collected at fraction no. 7.3 for 99mTc-cefprozil
which was found to coincide with the UV signal.

Figure 2. HPLC radiochromatogram of99mTc-cefprozil complex.

Factors affecting the labeling yield

Effect of ligand concentration

As shown in Figure 3, relatively low labeling yield of 99mTc-cefprozil, 75.3% ± 0.4%, was obtained at
low ligand concentration (1 mg). This low labeling yields was attributed to the ligand concentrations
being insufficient to form the complex with all of the reduced technetium-99m, while the percentage
of the colloid was high 14.9% ± 0.5%. Increasing the ligand concentration led to higher labeling
yield, and the maximum yield (97.5% ± 0.8%) was achieved at 4 mg. By increasing the ligand
concentration over the optimum values, the labeling yield remained stable.

23
Figure 3. Effect of cefprozil amount on the labeling yield of99mTc-cefprozil complex. Conditions:
1 to 10 mg of cefprozil, 50 μg Sn (II), pH 4, and 30-min reaction time; n = 3.

Effect of SnCl2·2H2O amount

The effect of the amount of stannous chloride was summarized in Figure 4. The data showed that the
radiochemical yield was dependent on the amount of SnCl2·2H2O present in the reaction mixture. At
25 μg SnCl2·2H2O, the labeling yield of 99mTc-cefprozil was 65.6% ± 0.71% due to insufficient
SnCl2·2H2O concentration to reduce all pertechnetate, so the percentage of was relatively high
(Guarna et al. [2001]). The labeling yield was significantly increased by increasing the amount of
SnCl2·2H2O from 25 to 200. At 50 μg (optimum content), the maximum labeling yield of
97.5% ± 0.8% was obtained. By increasing the amount of SnCl2·2H2O above the optimum
concentration value, the labeling yield decreased gradually due to the conversion of the excess
SnCl2·2H2O to colloid and reached to 55.4% ± 0.82% at 200 μg SnCl2·2H2O (Seung et al. [2002]).

Figure 4. Effect of Sn (II) amount on the labeling yield of99mTc-cefprozil complex. Conditions:
4 mg cefprozil, 25 to 200 μg Sn (II), pH 4, and 30-min reaction time; n = 3.

Effect of pH of the reaction mixture

24
As shown in Figure 5, at pH 1 the labeling yield of 99mTc-cefprozil complex was relatively low
(50%). The yield increased with increasing pH of the reaction mixture, reaching the maximum
labeling yield of 97.5% ± 0.8% at pH 4. By increasing the pH greater than 4, the labeling yield
decreased again to 62.8% at pH 6, at which the colloid impurity percentage becomes high (32.7% ±
0.3%) (Motaleb et al. [2012,2011]).

Figure 5. Effect of pH on the labeling yield of99mTc-cefprozil complex. Conditions: 4 mg

cefprozil, 50 μg Sn (II), pH 1 to 6, and 30-min reaction time; n = 3.

Effect of reaction time

Figure 6 shows the effect of incubation time on the radiochemical purity of the 99mTc-cefprozil
complex. At 1-min post labeling, the radiochemical purity reached to 80.6% ± 0.7%, which increased
with time until reaching its maximum value of 97.5% ± 0.8% at 30 min. The radiochemical purity
remains stable for up to 2 h (Ibrahim and Sanad [2013]; Sanad and Ibrahim [2013]).

Figure 6. Effect of reaction time on the labeling yield of99mTc-cefprozil complex. Conditions:
4 mg cefprozil, 50 μg Sn (II), pH = 4, 1- to 120-min reaction time; n = 3.

Stability test

25
As shown in Figure 7, the in vitro stability of 99mTc-cefprozil was studied in order to determine the
suitable time for injection to avoid the formation of the undesired products that result from the
decomposition of the complex. These undesired radioactive products might be accumulated in non-
target organs. The results of the stability test showed that the 99mTc-cefprozil is stable for 24 h at
37°C which resulted in a small release of radioactivity of 17.0% ± 0.3% (n = 3 experiments) from the
99m
Tc-cefprozil, as determined by paper chromatography.

Figure 7. In vitro stability of 99mTc- cefprozil in normal serum at optimum condition.

In vitro binding studies

In vitro binding studies revealed that the binding of 99mTc-cefprozil to S. aureus bacteria was similar
to that of 99mTc-ciprofloxacin, where the binding of 99mTc-cefprozil was in the range from 42% to
73% (n = 5), while the binding of 99mTc-ciprofloxacin was in the range from 40% to 65% (n = 3). The
varying amounts of cefprozil added (10- to 100-fold) showed significant decrease in the binding of
99m
Tc-cefprozil to living bacteria, indicating that the 99mTc-cefprozil complex is a specific agent for
bacterial cells (Figure 8).

Figure 8. In vitrobinding of99mTc-cefprozil toStaphylococcus aureus.

Biodistribution of 99mTc-cefprozil

26
Table 1 shows the biodistribution of 99mTc-cefprozil in important body organs and fluids. 99mTc-
cefprozil was removed from the circulation mainly through the kidneys and urine (approximately
42.79% injected dose, ID, at 4 h after injection of the tracer). The liver uptake decreased markedly
with time from 15.33% at 15 min to 6.9% at 4 h. The mice with infectious lesions injected with
99m
Tc-cefprozil showed a mean target-to-non-target (T/NT) ratio equal to 5.5 ± 0.12 which is greater
than that of 99mTc-ciprofloxacin (T/NT = 3.8 ± 0.8) (Ibrahim et al. [2011]). The accumulation of
activity at the site of infection was maximized at 2 h after intravenous injection then slightly
decreased with time until T/NT was equal to 3.70 ± 0.03 at 4 h post injection. During the first 2 h, the
radioactivity level in the blood was higher than that in the inflammation-induced area. This means
that the images of 99mTc-cefprozil by gamma camera will be affected by the blood pool images that
arise from the high blood activity. Therefore, it is recommended to obtain more specific images for
99m
Tc-cefprozil after 4 h (Figure 9).

Table 1. Biodistribution of99mTc-cefprozil inStaphylococcus aureusand turpentine oil-inflamed

mice at different time intervals

Figure 9. T/NT of99mTc-cefprozil complex in different inflammation models at different post

injection times.

Conclusion
99m
Tc-cefprozil was labeled easily using 50 μg SnCl2·2H2O as a reducing agent at pH 4 and 30-min
reaction time with high labeling yield of 97.5% ± 0.8%. The 99mTc-cefprozil complex was stable up
to 6 h, which shows high stability time in comparison to ciprofloxacin. Based on the data obtained
from the biodistribution of 99mTc-cefprozil, it can be stated that there is a significant difference in the
percentage uptake of 99mTc-cefprozil in the muscle injected with S. aureus and turpentine oil, which
indicated that 99mTc-cefprozil is considered as a specific agent and could distinguish between
bacterial infection and sterile inflammation. This result was supported by the data from in vitro
binding of 99mTc-cefprozil with bacteria.

Competing interests

The authors declare that they have no competing interests.

27
Authors’ contributions

EHB Make the analtycal parts. MHS makes the rest of manuscript. Both authors read and approved
the final manuscript.

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29
ABSTRACT EXAMPLES
Writing an Abstract – Examples
Each student who completes a science fair project must write an abstract to be displayed with the
project. An abstract gives the essence of the project in a brief but complete form — it should not
exceed 250 words. Judges and the public should have a fairly accurate idea of the project after
reading the abstract.
The abstract must focus on the current year’s research and give only minimal reference to previous
work. Details and discussions should not be included in the abstract, but may be put in the longer,
written research paper, or given on the project exhibit board.
Note that an abstract does not include acknowledgements (such as referencing mentor or university
laboratory) or a bibliography (this should be included in the Form 1A Research Plan Attachment).
The following colors in the two abstract examples demonstrate the following concepts:

Purpose of the Experiment (Blue)

• An introductory statement of the reason for investigating the topic of the project.
• A statement of the problem or hypothesis being studied.
Procedures Used (Green)
• A summarization of the key points and an overview of how the investigation was conducted.
• An abstract does not give details about the materials used unless it greatly influenced the procedure or
had to be developed to do the investigation.
• An abstract should only include procedures done by the student. Work done by a mentor (such as
surgical procedures) or work done prior to student involvement must not be included.
Observation/Data/Results (Red)
• This section should provide key results that lead directly to the conclusions you have drawn.
• It should not give too many details about the results nor include tables or graphs.
Conclusions (Pink)
• Conclusions from the investigation should be described briefly.
• The summary paragraph should reflect on the process and possibly state some applications and
extensions of the investigation.

JUNIOR HIGH LEVEL SAMPLE ABSTRACT

Sample Abstract Do Vitamin A Tablets Affect Plants
Jones, Mary E. 123 Main St., Hometown, IA 50001 Hometown Middle School, Hometown, IA
The purpose of this project was to determine if Vitamin A tablet have any effect on tomato plants. A total of
twelve Rutgers tomato plants each two inches tall were planted in identical individual plastic pots using two
cups of potting soil. Each plant received the same amount of water and sunlight during the three week
experiment. The twelve plants were divided into four groups of three plants each. One vitamin A tablet was
added to each of the three plants in the first group by burying the tablet one inch from the stem and one inch
deep. Two vitamin A tablets were added to the second group of three plants in a similar manner. The third
group of three plants had three tablets planted in the soil. The fourth group of three plants had no vitamin A
tablets added to the soil and served as the control group. The height of each plant was measured and
recorded at the start of the experiment and every 7 days thereafter. At the end of the experiment (21 days) the
stems were cut across at a height of 3 inches. Experimental groups showed less development and slower
growth rates than plants in the control group. The data was analyzed and the conclusion was drawn that giving
vitamin A tablets to tomato plants did not improve growth as each of the three experimental groups failed to
produce plants that were taller or had thicker stems than those in the control group

30
SENIOR HIGH LEVEL SAMPLE ABSTRACT

Sample Abstract Effects of Marine Exhaust Water on Algae

Jones, Sally C. 123 Main St., Hometown, IA 50001 Hometown High School, Hometown, IA
This project in its present form is the result of bioassay experimentation on the effects of two-cycle marine
engine exhaust water on certain green algae. The initial idea was to determine the toxicity of outboard engine
lubricant. Some success with lubricants eventually led to the formulation of “synthetic” exhaust water which, in
turn, led to the use of actual two-cycle engine exhaust water as the test substance.

Toxicity was determined by means of the standard bottle or “batch” bioassay technique. Scenedesmus
qaudricauda and Ankistrodesmus sp. Were used as the test organisms. Toxicity was measured in terms of a
decrease in the maximum standing crop. The effective concentration – 50 % (EC 50) for Scenedesmus
quadricauda was found to be 3.75% exhaust water, for Ankistrodesmus sp. 3.1% exhaust water using the
bottle technique.

Anomolies in growth curves raised the suspicion that evaporation was affecting the results; therefore, a flow-
through system was improvised utilizing the characteristics of a device called a Biomonitor. Use of the
Biomonitor lessened the influence of evaporation, and the EC 50 was found to be 1.4% exhaust water using
Ankistrodesmus sp. as the test organism. Mixed populations of various algae gave an EC 50 of 1.28% exhaust
water.
The contributions of this project are twofold. First, the toxicity of two-cycle marine engine exhaust was found to
be considerably greater than reported in the literature (1.4% vs 4.2%). Secondly, the benefits of a flow-through
bioassay technique utilizing the Biomonitor was demonstrated.

31